Recent findings suggest that regulatory interactions occur between the neural and skeletal systems. Using DNA microarrays, we found that expression of the serotonin transporter (5-HTT) is strongly up-regulated in RANKL-induced osteoclasts (OC). Fluoxetine ('Prozac'), a selective 5-HTT inhibitor, dose-dependently inhibited OC formation and resorptive activity. Mice homozygous for a deletion of the 5-htt gene exhibited increased cortical bone mass, and fluoxetine treatment of Swiss-Webster mice increased trabecular bone. These findings lead to the novel hypothesis that the serotonin (5-HT) system plays a crucial role in bone formation and remodeling, via effects on osteoclast development and/or activation. Aim 1 will characterize the expression of elements of the serotonin system in osteoclasts (OC) at the mRNA and protein levels, and will confirm that each is functional. Aim 2 will determine the role of serotonin (5-HT), the 5-HTT, serotonin receptors (SIR), vesicular monoamine transporter 1 (VMAT1), and monoamine oxidase (MAO) A in the formation and/or activation of OC in vitro, using a panel of specific agonists and inhibitors. In Aim 3, we will test the hypothesis that elevations in intracellular 5-HT are necessary for the observed effects, via effects on RANK-stimulated NFkB activation. Aim 4 will analyze the roles of the 5-HTT, SIRs, VMAT1, and MAO A in osteoclastogenesis in vivo using appropriate knockout mice. Aim 5 will determine the function of these serotonin system components in models of pathologic bone resorption, including resorption caused by estrogen-depletion or infection. The goal is to characterize the elements of the serotonin system that are present in OC, and determine their role in the regulation of bone mass under physiologic and pathologic conditions.